Engineered polypeptides are used widely in therapeutic and diagnostic applications. Therapeutic antibodies have been used for many years in, for example, treatment of cancer and inflammatory conditions. Therapeutic polypeptides are also used to treat and prevent blood conditions and viral infections. Diagnostic polypeptides have been used successfully to identify healthy and diseased cells and tissues in vivo.
Many polypeptides can provide targeting functionality to specific cells. The selective affinity of certain polypeptides can be used to target nearly any cell or tissue desired, for example a cell expressing an antigen. A polypeptide can carry a payload to destroy the target cell or tissue, or to slow its growth. Polypeptides have thus found use in therapy for conditions such as cancer, inflammatory diseases, autoimmune diseases and transplant rejection.
In certain applications therapeutic polypeptides are linked to molecular shields (e.g., macromolecules) to increase their half-life within an organism. For example, the attachment of polyethylene glycol (PEG) to a polypeptide can render the polypeptide non-detectable or less detectable by a patient's immune system, thereby reducing immunogenicity.
Polypeptides have also found use as diagnostics. These polypeptides can carry a label whose detection indicates the presence of a target receptor or antigen on a cell or in a tissue. These labels are typically linked to the polypeptides by covalent bonds.
To date, techniques for linking polypeptides to payloads such as molecular shields, labels, diagnostic compounds, and therapeutic compounds have been limited by their heterogeneity in degree and location of linking to the polypeptides, by their low yields and by losses in activity. These problems are particularly acute when attempting to conjugate more than one payload to a single polypeptide in a controlled manner, to produce a homogeneous product. Typical conjugation sites include random locations on polypeptide chains, e.g. random amines on amino acid residue side chains, and the N-terminus of certain polypeptide chains. In such cases, some polypeptides might be linked to a payload at one location while some polypeptides are linked to the same payload at another location, and some polypeptides might not be linked at all. If more than one payload is used, some polypeptides may be linked to a single payload, some polypeptides may be linked to all payloads, and some polypeptides may be linked to fewer than all payloads.
There is a need, therefore, for polypeptides modified at site-specific positions optimized for uniformity, yield and/or activity to further the promising use of polypeptides in, for example, therapy and diagnostics.